This is a competing renewal application to continue studies on the role of sodium- hydrogen exchanger 3 (NHE3) in the gastrointestinal tract. NHE belong to a family of transport proteins involved in the electroneutral exchange of Na+ and H+. NHE3 is the predominant Na+/H+ exchanger responsible for the majority of electrolyte transport across the gastrointestinal tract, and its inhibition by inflammatory mediators and enteropathogenic bacteria is believed to be the primary mechanism of inflammation- associated diarrhea. However, our recently published and preliminary data have demonstrated novel roles for NHE3 in the GI tract. These involve its contribution to epithelial integrity, modifying immune responses in intestinal inflammation, and in shaping the intestinal microbiota and its interactions with the mucosa. NHE3 deficiency leads to increased mucosal bacterial adherence and translocation, and to profound IBD- like dysbiosis. On the other hand, antibiotics and rederivation into a Helicobacter-free ultraclean barrier environment eliminates the inflammatory phenotype. Moreover, NHE3 deficiency results to impaired cellular adhesion, although the resulting barrier defect is partially compensated by regulatory immune responses in the intestinal mucosa. The current proposal is aimed at expanding upon these observations to address the following hypothesis: intestinal NHE3-mediated Na+/H+ exchange contributes to the intestinal homeostasis via modulating cellular interactions and colonic microbiota. Three specific aims have been formulated to address this hypothesis: (1) to determine the relationship between NHE3 and intestinal microbiota; (2) to determine the role of NHE3 in intestinal cell adhesion; and (3) to determine the role of mucosal regulatory immune responses in the compensation of epithelial defect in NHE3-/- mice. The proposed research will help explain the mechanisms by which NHE3 inhibition determines the extent of the epithelial barrier defect and contributes to the ultimate degree of inflammation, disease progression and outcome. PUBLIC HEALTH RELEVANCE: NHE3 is the predominant Na+/H+ exchanger responsible for the majority of sodium and water transport across the gastrointestinal tract, and its inhibition during inflammation represents the primary mechanism of inflammation-associated diarrhea. Novel data from our laboratory suggest additional consequences resulting from NHE3 inhibition, which may lead to profound alteration of intestinal microbiota, cellular interactions, and contribute to the ultimate epithelial barrier defect. The proposed studies are of significant basic and clinical relevance as they may not only explain the role of Na+/H+ exchange during intestinal inflammation but they may also guide future clinical approaches to a variety of inflammatory conditions of the gut.